Over the past several years, Kong and his colleagues have run dozens of experiments to find the right way to modify E2enough that the protein doesn't aggregate so readily and also so that the antibody-binding sites are maintained. This would enable the protein to be soluble and pure enough to grow crystals to determine its structure by the technique known as X-ray crystallography. "It was a Herculean effort," said Ward. "This is one of the most difficult and unstable viral envelope proteins around."
In the end, the team succeeded, using a slightly altered version of E2the E2 corewith some of its glycans (sugar molecules) and outer variable and stalk segments removed. The scientists were then able to obtain the high-resolution structure of the protein while it was bound to a known broadly neutralizing antibody developed at TSRI. The scientists then followed up by imaging a more complete version of E2 using electron microscopy to extend the structural model.
When finally revealed, E2's structure surprised the researchers. "It had been thought that HCV's E2 belongs to a family of viral fusion proteins called class 2 fusion proteins, which includes envelope proteins for West Nile and dengue viruses, for example," said Kong. "But we showed that E2 is structurally distinct and probably works differently than what had been widely assumed."
Based on the new structural data, Law and colleagues at TSRI are already designing and testing novel antibody-stimulating components of a future HCV vaccine. "Having the E2 structure has certainly helped us," said Law.
|Contact: Mika Ono|
Scripps Research Institute